Stem cells are primal cells that are found in all multi-cellular organisms. Stem cells have the ability to self-renew through mitotic cell division and can differentiate into a diverse range of specialized cell types. The three broad categories of mammalian stem cells are embryonic stem cells, adult stem cells, and cord blood stem cells. ES cells are derived from the inner cell mass of pre-implantation embryos, i.e., blastocysts. Human embryos reach the blastocyst stage 4-5 days post fertilization, at which time they consist of 50-150 cells. Adult stem cells are found in adult tissue. Cord blood stem cells are derived from the umbilical cord, which is rich in hematopoietic stem cells, i.e., stem cells that will form all cellular components of blood.
ES cells are pluripotent, meaning that they are able to differentiate into all of the somatic cell types of the three primary germ layers: the ectoderm, the endoderm, and the mesoderm. ES cells are the ultimate source for each of the more than 220 cell types in the adult body. When given no stimuli for differentiation, i.e. when grown in vitro, ES cells have the potential for indefinite expansion with continued plutipotency throughout each cell division.
Like embryonic stem cells, adult stem cells have the potential to self-renew indefinitely; however, unlike embryonic stem cells, adult stem cells are not pluripotent, rather, they are multipotent, meaning that they are able to differentiate into some, but not all specialized cells. Closely related to adult stem cells are adult progenitor cells. Like adult stem cells, adult progenitor cells are multipotent; however, unlike adult stem cells, adult progenitor cells cannot renew indefinitely. Having limited self-renewal abilities, adult progenitors differentiate into mature cells after several rounds of cell division. Examples of adult progenitors are satellite cells found in muscle and neural stem progenitor cells. Both adult stem cells and adult progenitors are found throughout the adult body and act as repair system for the organism by repopulating a limited number of cells types in the organism.
As stem cells can be readily grown and transformed in vitro into specialized cells with characteristics consistent with cells of various tissues, their use in medical therapies is of great interest. In order to effectively treat neural conditions with embryonic stem cells, there is a need in the art for an effective and efficient method to produce a homogeneous population of neural stem cells from embryonic stem cells. The following patent documents describe the isolation of neural stem cells from ES and adult progenitor cells.
U.S. Pat. No. 7,011,828 and U.S. Patent Publication Nos. 2005/0260747 A1 and 2006/0078543 A1 all to Reubinoff et al. teach the proliferation of an enriched population of embryonic stem cells, which are induced to differentiate in vitro to neural progenitor cells, neurons, and/or glial cells (collectively referred to as “neural cells”).
U.S. Pat. No. 7,015,037 and U.S. Patent Publication No. 2006/0030041 A1 both to Furcht et al. teach the differentiation of multipotent adult stem cells (MASCs) to form glial, neuronal, or oligodendrocyte cell types using growth factors, chemokines, and cytokines such as EGF, PDGF-BB, FGF2, and FGF-9.
U.S. Patent Publication No. 2006/0252149 to Zeigler teaches the maintenance of central nervous system (CNS) cells outside of an organism that retain the ability to proliferate and remain in a state of being able to differentiate as a result of exposure in culture to soluble laminin alone or together with one or more laminin associated factors (LAFs) and one or more of the CNS mitogens EGF, bFGF (also referred to as FGF2), and LIF.
U.S. Pat. No. 6,887,706 to Zhang et al. teaches a method of differentiating embryonic stem cells into neural precursor cells using the growth factor FGF2. In vitro differentiation of the ES cell-derived neural precursors was induced by withdrawal of FGF2 and plating on ornithine and laminin substrate.
U.S. Patent Publication No. 2007/0059823 A1 to Verfaillie et al. teaches a method for inducing ES cells and MASCs to differentiate into neuronal cells by culturing the stem cells initially with bFGF and later with FGF8, Sonic Hedgehog, brain-derived neutrotrophic factor, and astrocytes. Verfaillie et al. disclose that where it is desired for the stem cells to remain in an undifferentiated state, the media can contain supplements such as EGF, platelet derived growth factor (PDGF), and LIF.
U.S. Patent Publication No. 2005/0214941 A1 to Bhonsale et al. teaches a method for improving the growth rate of human fetal brain stem cells by culturing human neural stem cells (“hNSCs”) with bFGF, EGF, and LIF. Bhonsale et al. also provide a method for increasing the rate of proliferation of neural stem cell cultures and increasing the number of neurons in the differentiated cell population by culturing the hNSCs on a surface coated with polyornithine and fibronectin.
None of the foregoing patents and/or patent publications teaches the proliferation of a homogeneous population of neural stem cells with unlimited self-renewal capability derived from embryonic stem cells; accordingly, there remains a need in the art for an effective and efficient method of producing an unlimited supply of a homogeneous population of neural cells from embryonic stem cells.